Any feedback?
Please rate this page
(all_enzymes.php)
(0/150)

BRENDA support

5.3.3.1: steroid DELTA-isomerase

This is an abbreviated version!
For detailed information about steroid DELTA-isomerase, go to the full flat file.

Word Map on EC 5.3.3.1

Reaction

a 3-oxo-DELTA5-steroid
=
a 3-oxo-DELTA4-steroid

Synonyms

3-Keto-DELTA5-steroid isomerase, 3-Ketosteroid DELTA5-->DELTA4-isomerase, 3-ketosteroid isomerase, 3-Oxo steroid DELTA4-DELTA5-isomerase, 3-Oxo-delta5 steroid isomerase, 3-oxo-DELTA5-steroid isomerase, 3-Oxosteroid DELTA4-DELTA5-isomerase, 3-Oxosteroid DELTA5-DELTA4-isomerase, 3-Oxosteroid isomerase, 3beta-HSD, 3beta-HSD/isomerase, 3beta-hydroxysteroid dehydrogenase/5-ene-4-ene isomerase, 3beta-hydroxysteroid dehydrogenase/DELTA5-DELTA4 isomerase, 3beta-hydroxysteroid dehydrogenase/isomerase, 3beta-hydroxysteroid dehydrogenase/isomerase type 1, 3beta-hydroxysteroid dehydrogenase/isomerase type 2, 3KSI, 5-Ene-4-ene isomerase, 5-Pregnene-3,20-dione isomerase, delta 5-3-ketosteroid isomerase, DELTA-3-ketosteroid isomerase, Delta-5-3-ketosteroid isomerase, DELTA5(or DELTA4)-3-keto steroid isomerase, DELTA5-3-keto steroid isomerase, DELTA5-3-ketosteroid isomerase, DELTA5-3-oxosteroid isomerase, DELTA5-ketosteroid isomerase, DELTA5-steroid isomerase, glutathione transferase A3-3, GST A3-3, Hydroxysteroid isomerase, Isomerase, steroid DELTA, ketosteroid isomerase, KSI, More, Steroid 5-->4-isomerase, Steroid isomerase, TI, TI-WT, type I 3beta-hydroxysteroid dehydrogenase/isomerase

ECTree

     5 Isomerases
         5.3 Intramolecular oxidoreductases
             5.3.3 Transposing C=C bonds
                5.3.3.1 steroid DELTA-isomerase

Engineering

Engineering on EC 5.3.3.1 - steroid DELTA-isomerase

Please wait a moment until all data is loaded. This message will disappear when all data is loaded.
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D103A
-
site-directed mutagenesis of the catalytic residue, the mutant shows 5000fold reduced activity compared tot he wild-type enzyme
D38A
-
the catalytic turnover number is 140fold less than that for the wild-type
D38A/D99N
-
no detectable activity
D38A/Y14F
-
no detectable activity
D38A/Y55F
-
no detectable activity
D40N
-
site-directed mutagenesis, the mutation mimics the protonated aspartate found in the intermediate and equilenin complexes and leads to tighter binding of phenolate and other intermediate analogs
D99A
-
secondary kinetic isotope effects similar to wild-type. Mutation does not significantly decrease the contribution of coupled motion/hydrogen tunneling to the enzymatic reaction
D99L
the mutant shows about 10000fold decreased kcat compared to the wild type enzyme
F116W
the turnover-number for 5-androstene-3,17-dione is lowered 4.42fold, the KM-value is 3.1fold lower than the Km-value of the wild-type enzyme
Y14F/D99L
the mutant shows 1000000fold decreased kcat compared to the wild type enzyme
Y16F
-
site-directed mutagenesis of the catalytic residue, the mutant shows 50000fold reduced activity compared tot he wild-type enzyme
Y55F/Y88F
-
replacement of Tyr14 by 3-fluorotyrosine in the Y55,88F modified form of the isomerase results in a 4-fold decrease in turnover number
Y57F/D40N
-
site-directed mutagenesis, 3,4,5-trifluorophenol bind as ionized phenolate to KSI containing the Y57F mutation
D241N
-
mutant enzyme nearly has full isomerase activity, substantial decrease in 3beta-hydroxysteroid dehydrogenase activity. Mutant enzyme has a basal isomerase activity in the absence of coenzyme that is 10% of the NADH-stimulated turnover number
D257L
-
complete lack of isomerase activity in absence of coenzyme, substantial decrease in 3beta-hydroxysteroid dehydrogenase activity
D258L
-
complete lack of isomerase activity in absence of coenzyme, substantial decrease in 3beta-hydroxysteroid dehydrogenase activity
D265N
-
complete lack of isomerase activity in absence of coenzyme, substantial decrease in 3beta-hydroxysteroid dehydrogenase activity
D36A/K37R
-
mutation shifts the cofactor preference of both 3-beta-hydroxysteroid dehydrogenase and isomerase from NAD(H) to NADP(H)
DELTA283-310
-
the molecular weight of the subunit is 38800 Da compared to 42000 Da for the wild-type enzyme
E126L
site-directed mutagenesis, altered kinetics compared to the wild-type enzyme
H156Y
site-directed mutagenesis, leads to destabilization of interactions at the dimer interface and a dramatic increase in the substrate Km and inhibitor Ki values of 3beta-HSD isozyme 1 to equal those measured for 3beta-HSD isozyme 2
H232A
site-directed mutagenesis, altered kinetics compared to the wild-type enzyme
K158Q
site-directed mutagenesis, results in the complete loss of dehydrogenase activity and a reduction in isomerase activity, probably due to partial destabilization of cofactor binding, in which the catalytic triad is actively involved, and associated substrate binding
M187T
-
increase in affinity for NADH as an allosteric activator
N100A
site-directed mutagenesis, altered kinetics compared to the wild-type enzyme
N100S
site-directed mutagenesis, altered kinetics compared to the wild-type enzyme
N323L
site-directed mutagenesis, altered kinetics compared to the wild-type enzyme
Q105M
site-directed mutagenesis, leads to destabilization of interactions at the dimer interface and a dramatic increase in the substrate Km and inhibitor Ki values of 3beta-HSD isozyme 1 to equal those measured for 3beta-HSD isozyme 2
S124A
site-directed mutagenesis, results in the complete loss of dehydrogenase activity and a reduction in isomerase activity, probably due to partial destabilization of cofactor binding, in which the catalytic triad is actively involved, and associated substrate binding
S124T
-
11fold decrease in affinity to substrate
S322A
site-directed mutagenesis, altered kinetics compared to the wild-type enzyme
Y154F
site-directed mutagenesis, results in the complete loss of dehydrogenase activity and a reduction in isomerase activity, probably due to partial destabilization of cofactor binding, in which the catalytic triad is actively involved, and associated substrate binding
D103N/D40N
site-directed mutagenesis, 3,4,5-trifluorophenol bind as ionized phenolate to KSI containing the D103N mutation
D99E
-
turnover-number for 5-androstene-3,17-dione is 68.1% of the turnover number of the wild-type enzyme, the Km-value is 1.22fold higher than the Km-value of the wild-type enzyme
E118A
-
the free-energy change for unfolding in the absence of urea at 25°C is decreased by about 3.9 kcal/mol compared to wild-type value. Mutation increasex the dissociation constant for (+)-equilenin, a reaction intermediate analogue. 50% of the protein is unfolded at 4.46 M urea compared to 5.22 M for the wild-type enzyme. The turnover-number for 5-androstene-3,17-dione is 33% of that of the wild-type enzyme, the Km-value is 348% of that of the wild-type enzyme
E118A/N120A
-
the free-energy change for unfolding in the absence of urea at 25°C is decreased by about 9.5 kcal/mol compared to wild-type value. Mutation increases the dissociation constant for (+)-equilenin, a reaction intermediate analogue. 50% of the protein is unfolded at 3.89 M urea compared to 5.22 M for the wild-type enzyme. The turnover-number for 5-androstene-3,17-dione is 6% of that of the wild-type enzyme, the Km-value is 523% of that of the wild-type enzyme
F54A
-
turnover number for 5-androstene-3,17-dione is 17.1% of the turnover number for the wild-type enzyme, turnover number for 5(10)-estrene-3,17-dione is 15.3% of the turnover number of the wild-type enzyme,the KM-value for 5-androstene-3,17-dione is 116% of the KM-value of the wild-type enzyme, the KM-value for 5(10)-estrene-3,17-dione is 233% of the KM-value of the wild-type enzyme
F54L
-
turnover number for 5-androstene-3,17-dione is 41.9% of the turnover number of the wild-type enzyme, turnover number for 5(10)-estrene-3,17-dione is 38.7% of the turnover number of the wild-type enzyme, the KM-value for 5-androstene-3,17-dione is 154% of the KM-value of the wild-type enzyme, the KM-value for 5(10)-estrene-3,17-dione is 89.3% of the KM-value of the wild-type enzyme
F82A
-
turnover number for 5-androstene-3,17-dione is 2.5% of the turnover number of the wild-type enzyme, turnover number for 5(10)-estrene-3,17-dione is 2.3% of the turnover number of the wild-type enzyme,the KM-value for 5-androstene-3,17-dione is 29.9% of the KM-value of the wild-type enzyme, the KM-value for 5(10)-estrene-3,17-dione is 29.8% of the KM-value of the wild-type enzyme
F82L
-
turnover number for 5-androstene-3,17-dione is 12% of the turnover number of the wild-type enzyme, turnover number for 5(10)-estrene-3,17-dione is 67.6% of the turnover number of the wild-type enzyme, the KM-value for 5-androstene-3,17-dione is 108% of the KM-value of the wild-type enzyme, the KM-value for 5(10)-estrene-3,17-dione is 130% of the KM-value of the wild-type enzyme
L125A/V127A
-
mutation in small exterior hydrophobic cluster, decrease in conformational stability
L125F/V127F
-
mutation in small exterior hydrophobic cluster, slight increase in stability
N120A
-
the free-energy change for unfolding in the absence of urea at 25°C is decreased by about 7.8 kcal/mol compared to wild-type value. Mutation increasex the dissociation constant for (+)-equilenin, a reaction intermediate analogue. 50% of the protein is unfolded at 3.95 M urea compared to 5.22 M for the wild-type enzyme. The turnover-number for 5-androstene-3,17-dione is 17% of that of the wild-type enzyme, the Km-value is 516% of that of the wild-type enzyme
R72A
-
the free-energy change for unfolding in the absence of urea at 25°C is decreased by about 3.8 kcal/mol compared to wild-type value. Mutation increasex the dissociation constant for (+)-equilenin, a reaction intermediate analogue. 50% of the protein is unfolded at 4.74 M urea compared to 5.22 M for the wild-type enzyme. The turnover-number for 5-androstene-3,17-dione is 23% of that of the wild-type enzyme
W116A
-
turnover number for 5-androstene-3,17-dione is 1.5% of the turnover number of the wild-type enzyme, turnover number for 5(10)-estrene-3,17-dione is below 12%% of the turnover number of the wild-type enzyme, the KM-value for 5-androstene-3,17-dione is 348% of the KM-value of the wild-type enzyme
W116F
-
turnover number for 5-androstene-3,17-dione is 22.4% of the turnover number of the wild-type enzyme, the KM-value for 5(10)-estrene-3,17-dione is 299% of the KM-value of the wild-type enzyme
W116Y
-
turnover number for 5-androstene-3,17-dione is 21% of the turnover number of the wild-type enzyme
W92A
-
mutation in small exterior hydrophobic cluster, decrease in conformational stability. Crystallization data
W92A/L125A/V127A
-
mutation in small exterior hydrophobic cluster, decrease in conformational stability
Y115F
the mutant shows about 68% activity compared to the wild type enzyme
Y14F/D99E
-
turnover-number for 5-androstene-3,17-dione is less than 1% of the turnover number of the wild-type enzyme, the Km-value is 1.05fold lower than the Km-value of the wild-type enzyme
Y14F/D99L
Y14F/D99N
-
turnover-number for 5-androstene-3,17-dione is less than 1% of the turnover number of the wild-type enzyme, the Km-value is 1.06fold lower than the Km-value of the wild-type enzyme
Y14F/Y30F
-
the turnover-number for 5-androstene-3,17-dione is about 1% of that of the wild-type enzyme, the Km-value is 1.57fold higher than that of the wild-type enzyme
Y14F/Y30F/Y55F
-
the turnover-number for 5-androstene-3,17-dione is about 2.1% of that of the wild-type enzyme, the Km-value is comparable to the Km-value of the wild-type enzyme. The hydrogen bond between Asp99 Odelta2 and C3-O of the steroid, which is perturbed by the Y14F mutation, can be partially restored to that in the wild-type enzyme by additional Y30F/Y55F mutations. The improvement in the catalytic activity of Y14F by the additional Y30f/Y55F mutation is due to the changes in the structural integrity at the catalytic site and the resulting restoration of the proton-transfer mechanism in Y14F/Y30F/Y55F
Y14F/Y30F/Y55F/D99L
-
the turnover-number for 5-androstene-3,17-dione is less than 1.2% of that of the wild-type enzyme, the Km-value is 1.8fold higher than that of the wild-type enzyme
Y14F/Y30F/Y55F/D99N
-
the turnover-number for 5-androstene-3,17-dione is less than 1% of that of the wild-type enzyme, the Km-value is 1.14fold higher than that of the wild-type enzyme
Y14F/Y55F
-
the turnover-number for 5-androstene-3,17-dione is about 1.2% of that of the wild-type enzyme, the Km-value is 1.75fold higher than that of the wild-type enzyme
Y16F/Y32F/Y57F
the number of water molecules directly hydrogen bonded to the ligand oxygen is one in the Y16F/Y32F/Y57F mutant
Y16S
the number of water molecules directly hydrogen bonded to the ligand oxygen is approximately two in the Y16S mutant
Y30F/D99L
Y30F/Y55F
Y30F/Y55F/Y115F
the mutant shows about 57% activity compared to the wild type enzyme
Y55F/D99L
Y57S
-
the mutation causes a large decrease in the catalytic performance of the enzyme